European Journal of Clinical Investigation (2000) 30, 751±753 Paper 724

Commentary

Naturally occurring mutations of hepatitis B virus andoutcome of chronic infection: Is there an association?

S. GuÈnther

Bernhard-Nocht Institut fuÈr Tropenmedizin, Hamburg, Germany

See paper on page 787

A focus of hepatitis B virus (HBV) research was, and still is,

the study of the genetic variability of HBV. One major goal

behind the intensive efforts is to identify mutation(s) in the

HBV genome which codetermine the course or outcome of

infection, or the ef®ciency of antiviral therapy.

Although HBV is a DNA virus, it replicates via an RNA

intermediateÐthe RNA pregenomeÐwhich is reverse

transcribed into HBV DNA by the virus-encoded poly-

merase. It is generally assumed that this reverse transcrip-

tion step accounts for the majority of point mutations and

deletions or insertions that can be observed in the HBV

genome.

There are two major types of mutations in HBV. Firstly,

there are genotype-speci®c mutations which are stably

transmitted in the host population and allow the distinction

of currently six genotypes. These genotypes cluster geo-

graphically. Although the existence of genotypes is known

for a long period of time, only very recently an association

of genotype and clinical outcome was proposed [1,2]. The

second type of HBV variability concerns mutations that

emerge in an individual during chronic infection. Several

speci®c mutations of this type have been identi®ed by a

large number of longitudinal as well as cross-sectional

studies conducted during the past decade (for a recent

review see [3]).

Most of the corresponding variants accumulate during

infection and persist as a dominant population until late

phases. A recent study calculated that an average of 20

mutations affecting all virus genes accumulate in the domi-

nant virus population if infection takes place in childhood

and proceeds, over time, up to late stages [4]. Mutations

emerging during infection seem not to cluster geographi-

cally unless genotype-speci®c mutations interfere with their

introduction. Moreover, we learned that the presence or

emergence of speci®c mutations is associated with parti-

cular stages of chronic infection [3]. For example, A !T

and G ! A mutations at position 1762/1764 in the prege-

nomic promoter (T/A-1762/64 mutations) are character-

istic for patients with signs of liver in¯ammation (see below

for details).

Pre-C mutations (commonly a stop codon), which pre-

vent synthesis of hepatitis B e antigen (HBeAg) as well as

amino acid changes at speci®c positions in the nucleocapsid

gene, generally emerge later than the T/A-1762/64 muta-

tions during seroconversion from HBeAg to the corre-

sponding antibody (anti-HBe). Deletions in the pre-S2

region and mutations preventing synthesis of the pre-S2

protein are also characteristic of late stages of infection.

Mutations in the hepatitis B s antigen (HBsAg) `̀ a''

determinant-changing antigenicity of HBsAg, are charac-

teristic for the phase following loss of HBsAg and/or

appearance of antibodies to HBsAg. Accordingly, HBV

genomes from patients with long-standing infection who

developed cirrhosis or hepatocellular carcinoma (HCC)

have often accumulated these mutations [5]. However,

the same mutations are also seen in patients who went

into remission. Although in the past some studies have

claimed an in¯uence of speci®c mutations on the outcome

of hepatitis B, this could often not be substantiated by

others. A compilation of published clinical and sequence

data of thousands of patients could also not establish such

an association, in particular with respect to pre-C mutation

and amino acid changes in the nucleocapsid gene [3].

The current data are most consistent with the interpre-

tation that mutations introduced in the HBV genome

during natural infection, are predominantly a marker of

the duration or severity of liver in¯ammation, or type and

intensity of the immune response, rather than a determi-

nant of the outcome. Long-term follow up studies analys-

ing complete genomes are required to prove or disprove this

speculation.

A typical example of a mutation that is associated

with particular stages of chronic infection are the above

Q 2000 Blackwell Science Ltd

Bernhard-Nocht Institut fuÈr Tropenmedizin (S. GuÈnther)

Hamburg, Germany.

Correspondence: Stephan GuÈnther, Bernhard-Nocht Institut fuÈr

Tropenmedizin, Bernhard-Nocht Strasse 74, D-20359

Hamburg, Germany. Tel.: �49 40 42818 421; Fax: �49

40 42818 378; e-mail: guenther@bni.uni-hamburg.de

See paper on page 000

Received 15 June 2000; accepted 18 June 2000

752 S. GuÈnther

mentioned A !T and G ! A mutations at position 1762/

64 in the pregenomic promoter. These mutations are

observed in only 10% of patients in the ®rst asymptomatic

phase of chronic infection. This phase typically follows

perinatal infection and is characterized by high viremia,

HBeAg in the circulation, and low or absent in¯ammatory

liver disease (normal alanine aminotransferase [ALT]

levels). However, these mutations are found in about

60% of patients with in¯ammatory liver disease, irrespec-

tive of the HBeAg status. About 80% of patients who have

developed cirrhosis or HCC are infected by HBV charac-

terized by these mutations (the prevalence data are from a

compilation in [3]).

Why these variants become selected apparently with the

development of hepatitis is speculative. Enhanced replica-

tion, as observed in cell culture by some though not all

investigators [6±8], is one possibility. In addition, the

functional studies indicated that these mutations reduce

the expression of HBeAg on the transcriptional level, which

may suggest that the corresponding variants are selected by

the immune response or other mechanisms because they

produce less HBeAg. Since these mutations alter the bind-

ing of transcription factors to the core promoter [9,10] and

are closely associated with liver in¯ammation, it is also

conceivable that they adapt HBV to in¯ammation-

mediated changes in the composition of hepatic

transcription factors.

Singaporean researchers report in this issue of the

European Journal of Clinical Investigation cross-sectional

data suggesting that the association of liver disease with

the occurrence of T/A-1762/64 mutations is not as strong

as currently thought, at least in Singaporean HBV carriers

[11]. In their cohort of patients with chronic infection and

liver in¯ammation, as indicated by elevated ALT, they

found T/A-1762/64 mutations in only 15% of the patients.

However, in agreement with previous reports they observed

T/A-1762/64 mutations in 70% of their patients with

cirrhosis or HCC.

A possible interpretation of these results is that the

introduction of T/A-1762/64 mutations during the course

of infection is somewhat delayed in Singaporean HBV

carriers compared to patients in other geographic regions.

This view is supported by recent reports which suggest that

T/A-1762/64 mutations occur only rarely in East Asian

patients infected with particular HBV strains or genotypes

[2,12]. Provided a speci®c genotype that circulates in Asia

(genotype B) is indeed associated with a reduced level of

liver disease, as suggested by two recent studies [1,2], and

assuming that the immunological or pathophysiological

processes associated with this reduced level of disease

may not be suf®cient to drive selection of T/A-1762/64

variants, this could lead to a lower prevalence of these

mutations. Further research is clearly warranted to deter-

mine more precisely host and virus conditions leading to

selection of these variants.

Another interesting ®nding of the study [11] is the

exceptionally high viremia in the three patients infected

by HBV with deletions in the core promoter compared with

the viremia in patients infected with T/A-1762/64 variants.

This type of mutation has been rarely documented in the

available literature. Consistent with the present study, core

promoter deletions are usually found in patients who are

positive for HBeAg, have chronic infection with elevated

ALT, or have developed cirrhosis or HCC [3]. Experiments

with variants characterized by similar deletions indicated

slightly enhanced replication in cell culture [13,14],

although the deletions remove part of the core promoter

that drives the synthesis of the RNA pregenome.

Even though in vitro replication data can hardly be

extrapolated to the complex in vivo situation, the data

reported in the present study provide a ®rst clue that the

enhanced replication capacity of these variants in vitro may

indeed lead to higher viremia in vivo. Whether this is of any

relevance to the outcome of infection is speculative. Due to

the rare occurrence of these mutations, it will be dif®cult to

assess their epidemiology as well as clinical relevance.

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